Dasypyrum Breviaristatum
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Li et al. Molecular Cytogenetics (2016) 9:6 DOI 10.1186/s13039-016-0217-0 RESEARCH Open Access Molecular cytogenetic characterization of Dasypyrum breviaristatum chromosomes in wheat background revealing the genomic divergence between Dasypyrum species Guangrong Li, Dan Gao, Hongjun Zhang, Jianbo Li, Hongjin Wang, Shixiao La, jiwei Ma and Zujun Yang* Abstract Background: The uncultivated species Dasypyrum breviaristatum carries novel diseases resistance and agronomically important genes of potential use for wheat improvement. The development of new wheat-D. breviaristatum derivatives lines with disease resistance provides an opportunity for the identification and localization of resistance genes on specific Dasypyrum chromosomes. The comparison of wheat-D. breviaristatum derivatives to the wheat-D. villosum derivatives enables to reveal the genomic divergence between D. breviaristatum and D. villosum. Results: Themitoticmetaphaseofthewheat-D. breviaristatum partial amphiploid TDH-2 and durum wheat -D. villosum amphiploid TDV-1 were studied using multicolor fluorescent in situ hybridization (FISH). We found that the distribution of FISH signals of telomeric, subtelomeric and centromeric regions on the D. breviaristatum chromosomes was different from those of D. villosum chromosomes by the probes of Oligo-pSc119.2, Oligo-pTa535, Oligo-(GAA)7 and Oligo-pHv62-1. A wheat line D2139, selected from a cross between wheat lines MY11 and TDH-2, was characterized by FISH and PCR-based molecular markers. FISH analysis demonstrated that D2139 contained 44 chromosomes including a pair of D. breviaristatum chromosomes which had originated from the partial amphiploid TDH-2. Molecular markers confirmed that the introduced D. breviaristatum chromosomes belonged to homoeologous group 7, indicating that D2139 was a 7Vb disomic addition line. The D2139 displayed high resistance to wheat stripe rust races at adult stage plant, which may be inherited from, D. breviaristatum chromosome 7Vb. Conclusion: The study present here revealed that the large divergence between D. breviaristatum and D. villosum with respected to the organization of different repetitive sequences. The identified wheat- D. breviaristatum chromosome addition line D2139 will be used to produce agronomically desirable germplasm for wheat breeding. Keywords: Dasypyrum breviaristatum, Fluorescence in situ hybridization, Molecular marker, Wheat Background VVVbVb. Dasypyrum species possess agronomically im- The genus Dasypyrum (or Haynaldia) consists of only portant genes such as disease resistance, high protein two diploid species, the annual Dasypyrum villosum and quality and drought tolerance, all of which represent valu- perennial D. breviaristatum [1]. The genomes of diploid able resources for global wheat breeding [3]. The species D. villosum and D. breviaristatum were assigned the sym- D. villosum has been extensively hybridized with wheat for bols V and Vb, respectively [2, 3]. Based on the sequences at least 6 decades, and several disease resistance genes comparison of nr5S DNA multigene family, Baum et al. have been successfully transferred to wheat [5–7]. Above [4] suggested that the genome constitution of 4x D. all, over 20 elite cultivars carrying the wheat- D. villosum breviaristatum should be considered as an allotetraploid chromosome T6AL · 6VS translocation with powdery mildew resistance gene Pm21 have been released into agri- cultural production in China [8, 9]. Given the widespread * Correspondence: [email protected] School of Life Science and Technology, University of Electronic Science and success of this introgression from D. villosum, researches Technology of China, Chengdu 610054, China © 2016 Li et al. Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http:// creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Li et al. Molecular Cytogenetics (2016) 9:6 Page 2 of 9 have been conducted with a similar aim to transfer useful be related to the presence of D. breviaristatum chro- genes from D. breviaristatum into wheat. Subsequently, mosomes. Each of the seven pairs of D. breviaristatum the wheat- D. breviaristatum partial amphiploid [10] and chromosomes were also distinguished using probes Oligo- wheat- D. breviaristatum introgression lines with multiply pSc119.2, Oligo-pTa535 in TDH-2 (Fig. 1b). These chro- disease resistances have developed [11, 12]. mosomes were temporarily named Vb1-Vb7 (Fig. 1b). Molecular and cytogenetic methods have been previ- FISH using probes Oligo-pSc119.2, Oligo-pTa535, Oligo- ously employed to assess the level of chromosomal di- (GAA)7 were also carried out on the chromosomes of the vergence of these two Dasypyrum species [13, 14]. A Triticum turgidum cv. Jorc-69- D. villosum amphiploid large number of interspecific and intraspecific chromo- TDV-1(Fig.1candd).Wefoundthatthesignalsofprobe some variations and significant genomic diversification Oligo-pSc119.2 were mainly located on terminal sites, were observed among different Dasypyrum accessions, while the hybridization signals of Oligo-pTa535 were probably due to the out-crossing characteristic of these distributed along the chromosome arms of D. villosum. two Dasypyrum species [2]. Each pair of Dasypyrum The probe Oligo- (GAA)7 hybridized to 2V-7V of D. villo- chromosomes were transferred into a wheat background sum chromosomes at their centromeric or sub-terminal after intergeneric hybridizations. The wheat- Dasypyrum regions. Moreover, we produced a high tandem repeat chromosome addition lines with different Dasypyrum sequences probe Oligo-pHv62-1 as reported by Li et al. species or accession origins allow comparison of the [17]. FISH revealed that Oligo-pHv62-1 present in ter- different Dasypyrum genomes in wheat backgrounds. In minal or sub-terminal heterochromatic C-banding regions the present study, fluorescent in situ hybridization (FISH) of D. villosum chromosomes in TDV-1, but was absent in was carried out to characterize differences between D. bre- D. breviaristatum chromosomes of TDH-2 (Fig. 1e and f). viaristatum and D. villosum chromosomes by comparing The comparative FISH karyotypes of the D. breviarista- karyotypes between the wheat- D. breviaristatum partial tum and D. villosum chromosomes allows easily to distin- amphiploid TDH-2 and Triticum turgidum - D. villosum guish each individual Dasypyrum chromosome in wheat amphiploid TDV-1, The FISH and molecular markers background. were applied to identify new wheat- D. breviaristatum addition line with stripe rust resistance, which will be a FISH of wheat- D. breviaristatum addition line D2139 useful germplasm for wheat genetics and breeding. Sequential multi-color ND-FISH by probes Oligo-pSc119.2, Oligo-pTa535, Oligo-(GAA)7 was conducted to charac- Results terize the mitotic metaphase cells of D2139 (Fig. 2). Comparative FISH karyotype of TDH-2 and TDV-1 The chromosome number of D2139 is 2n = 44, including The mitotic metaphase chromosomes of the wheat- D. all the 42 wheat chromosomes and two alien chromo- breviaristatum partial amphiploid TDH-2, were hybridized somes added in the wheat background. The probes Oligo- with probes Oligo-pSc119.2, Oligo-pTa535, Oligo-(GAA)7 pSc119.2, Oligo-pTa535, showed a pair of chromosomes by sequential multicolor-FISH (Fig. 1). As shown in with faint Oligo-pSc119.2 hybridization signals at the telo- Fig. 1a, the FISH hybridization signals of the probes meric region of long arm, and strong hybridization signals Oligo-pSc119.2 and Oligo-pTa535 can easily identify of Oligo-pTa535 along the long and short arm in D2139 the 28 wheat chromosomes from 1A-7A and 1B-7B (Fig. 2). The FISH hybridization pattern of the chromo- based on the standard FISH karyotype of wheat chromo- somes was identical to D. breviaristatum chromosomes somes using the same probes described by Tang et al. [15]. Vb7 (Fig. 1). Therefore, we concluded that the line D2139 Yang et al. [10] reported that the partial amphiploid TDH-2 was a chromosome Vb7 addition line. Comparing the was produced by the elimination of some chromosomes FISH patterns of D2139 parents MY11 [15] and TDH-2 fromthewheatChineseSpring(CS)-D. breviaristatum (Fig. 1), it appeared that the D2139 line inherited the A decaploid amphiploid. It is likely that the A and B chro- and B- genome chromosomes from MY11 and/or TDH-2. mosomes of TDH-2 originated from CS. By comparing Based on the FISH patterns, D2139 inherited chromosomes the FISH patterns of Oligo-pSc119.2 and Oligo-pTa535 5A, 7A, 1B, and 7B which were identical to the TDH-2 par- probes of TDH-2 to those of CS by Tang et al. [15], we ent, while 6B, 2B appeared to be from MY11. Since there is found additional signals corresponding to probe Oligo- no D-genome in the partial amphiploid TDH-2, D2139 pSc119.2 on the terminal regions of 1BS and 2BL in would have inherited D-chromosomes from MY11. As TDH-2 (Fig. 1b). After comparing the (GAA)n signal shown in Fig. 2c, chromosomes 1D and 3D revealed clear distribution on TDH-2 chromosomes with those of CS differences in the distribution of Oligo-pSc119.2 signals reported by Danilova